Use of commercial somatic cell counters to quantify somatic cells in non-lactating bovine mammary gland secretions.

Measurement of the somatic cell count (SCC) in milk is commonly used to detect mastitis in lactating dairy cows. Many techniques and tools have been developed and adapted to quantify milk SCC, but few tools have been evaluated in their ability to enumerate somatic cells in non-lactating bovine mammary secretions. This limits the tools available for detecting mastitis in non-lactating animals. The objective of these studies was to evaluate methods of somatic cell quantification, originally developed for milk, in their ability to quantify the SCC in non-lactating bovine mammary secretions when compared to the gold standard microscopic quantification method. Two experiments were conducted. In a first experiment, 222 mammary secretions were collected and diluted 1:10 with PBS. Cells in these suspensions were quantified microscopically and with a DeLaval Cell Counter. Microscopic SCC (MSCC) ranged from 1.9 × 106 to 259.5 × 106 cells/mL while DeLaval Cell Counter SCC (DSCC) ranged from 1.8 × 106 to 27.0 × 106 cells/mL; a measurement of agreement between the 2 measures, based on the Lin's Concordance Correlation Coefficient (CCC) suggested moderate agreement between measures (CCC = 0.60). In a second experiment 72 mammary secretions were collected and diluted 1:50 in PBS. Somatic cells in these suspensions were quantified microscopically, with a DeLaval Cell Counter, and by a DHIA laboratory using a Fossomatic™ FC. MSCC ranged from 1.6 to 47.5 × 106 cells/mL, DSCC ranged from 1.0 to 35.7 × 106 cells/mL, and Fossomatic SCC (FMSCC) ranged from 1.6 to 46.7 × 106 cells/mL. CCCs of 0.81 and 0.88 resulted when DSCC and FMSCC were paired with the MSCC, respectively. The results of this work indicate that a significantly greater concentration of somatic cells exist in non-lactating mammary secretions and dilution of these mammary secretions influences accuracy of SCC estimates. Future studies seeking to quantify somatic cells in mammary secretions from non-lactating cows should identify the most appropriate dilution factors specific to each method of measure, given that these two factors will influence the accuracy of SCC estimates. Development of a standardized approach for quantifying somatic cells in non-lactating dairy animals such as heifers and cows, via a rapid automated counter, can allow for the detection of mastitis in non-lactating dairy animals.

An immunomodulatory feed additive enhances in vitro viral vaccine recall antigen responses in dairy heifers.

Enhancing immunological responses to vaccination is an important goal in many herd health management systems. OmniGen-AF®(OG) is an immunomodulatory feed additive that has been shown to enhance innate immune function in ruminants and its effects on adaptive immunity require additional study. The objective of this study was to evaluate post-vaccine antibody titers and circulating cellular memory development in heifers fed OG and administered a commercially available modified-live bovine respiratory disease (BRD) vaccine. Twenty-four Holstein heifers were assigned to one of two diets for 170 days: Control TMR (CON; n = 11), or TMR plus OG (TRT; 9 g/100 kg BW/day; n = 13). Samples for hematology, serology, and cellular assays were collected on D-110, 0, 21, 42, and 60 of the trial. Heifers were administered two priming doses of a modified-live BRD vaccine, with a third dose given on D0. There were no significant differences in total WBC and absolute number or the percentage of circulating lymphocytes, monocytes, neutrophils, RBC, or platelets on D-110 through D21. On D42 and D60, CON had significantly higher numbers of lymphocytes. On D0, mean serum neutralizing (SN) titer to BHV-1 was significantly higher for CON compared to TRT. SN titers were not significantly different between CON and TRT at any other time point for BHV-1, BVDV type 1, or BVDV type 2. TRT mounted a significantly stronger recall proliferative response to 0.5 multiplicity of infection (MOI) of BHV-1, BVDV type 1 and BVDV type 2 on D42 and D60; 0.25 MOI of BVDV type 1 on D21 and D42; and 0.25 MOI BVDV type 2 on D42 compared to CON. IL-4 production induced by 0.5 and 1.0 MOI BHV-1 (D42 and D60); 0.25 MOI of BVDV type 1 (D21); and 0.25 and 0.5 MOI of BVDV type 2 (D60) were significantly higher for TRT than CON. IL-17 production induced by 0.25 MOI of BVDV type 1 was significantly higher on D60 for TRT compared to CON. IFN-gamma and IL-10 were not significantly different between treatments. These data indicate feeding OG has a beneficial effect on responses to vaccine antigens in Holstein dairy heifers.

Effects of an immunomodulatory feed additive on intramammary infection prevalence and somatic cell counts in a dairy herd experiencing major health issues.

A trial was conducted to determine if feeding OmniGen-AF® (OG) to 22 late lactation cows 60 days prior to and during the early dry period, a time of increased susceptibility to mastitis, could reduce disease incidence in a dairy herd experiencing major health issues. Treated cows (n = 11) consumed a ration containing OG [9 g/100 kg of body weight/day] beginning 60 days before dry-off, during the dry period, and through 30 days in milk (DIM). Control cows received the same ration during the dry period through 30 DIM only. Body weights, body condition scores (BCS), intramammary infection (IMI) prevalence, new IMI rates, somatic cell counts (SCC), milk yield, and adverse health events were measured. No differences were found between treatments for body weight or BCS. Adverse health event data at calving showed no differences between treatments except for percentage of cows with hyperketonemia, which was lower among treated cows (63.6% vs 100%). Prevalence of IMI from calving through 30 DIM for treated cows (6.1%) was lower than controls (11.05%); likewise, new IMI rate during this time for treated cows (0.61%) was lower than controls (5.81%). The SCC from calving through 30 DIM for treated cows (215,000/ml) was lower than controls (493,000/ml). Average production/day at the first DHIA test (~33 DIM) showed that treated cows produced more milk (39.9 kg) than controls (35.34 kg). In conclusion, feeding OG 60 days prior to dry-off reduced hyperketonemia and mastitis, lowered SCC, and numerically increased milk yield in a dairy herd experiencing major health issues.

Effects of Staphylococcus aureus intramammary infection on the expression of estrogen receptor α and progesterone receptor in mammary glands of nonlactating cows administered estradiol and progesterone to stimulate mammary growth.

Intramammary infections (IMI) are prevalent in nonlactating dairy cattle and are known to alter mammary structure and negatively affect the amount of mammary epithelium in the gland. Mechanisms responsible for the observed changes in mammary growth during an IMI are poorly understood, yet the importance of the key mammogenic hormones driving mammary growth is well recognized. This study's objective was to characterize the expression of estrogen receptor α (ESR1) and progesterone receptor (PGR) in mammary glands stimulated to grow and develop in the presence or absence of an IMI as well as preliminarily characterize myoepithelial cell response to IMI. Mammary growth was stimulated in 18 nonpregnant, nonlactating dairy cows using subcutaneous estradiol and progesterone injections, and 2 culture-negative quarters of each cow were subsequently infused with either saline (n = 18) or Staphylococcus aureus (n = 18). Mammary parenchyma tissues were collected 5 d (n = 9) or 10 d (n = 9) postchallenge and examined using immunofluorescence microscopy to quantify positive nuclei and characterize staining features. There tended to be a greater number of ESR1-positive nuclei observed across 8 random mammary parenchyma fields of view in saline quarters than in Staph. aureus quarters (201 vs. 163 ± 44 nuclei). Saline quarters also contained a greater number of PGR-positive nuclei (520 vs. 440 ± 45 nuclei) and myoepithelial cells (971 vs. 863 ± 48 nuclei) than Staph. aureus-challenged quarters. However, when ESR1, PGR, and myoepithelial nuclei counts were adjusted for Staph. aureus quarters containing less epithelium, differences between quarter treatments abated. The examined ESR1 and PGR staining characteristics were similar between saline and Staph. aureus quarters but were differentially affected by day of tissue collection. Additionally, nuclear staining area of myoepithelial cells was greater in Staph. aureus quarters than in saline quarters. These results indicate that IMI had little effect on the number or staining characteristics of ESR1- or PGR-positive nuclei relative to epithelial area, but myoepithelial cells appear to be affected by IMI and the associated inflammation in nonlactating mammary glands that were stimulated to grow rapidly using mammogenic hormones. Accordingly, reductions in mammary epithelium in affected glands are not suspected to be resultant of alterations in the number or staining characteristics of ESR1- or PGR-positive mammary epithelial cells.

Apoptosis and proliferation in Staphylococcus aureus-challenged, nonlactating mammary glands stimulated to grow rapidly and develop with estradiol and progesterone.

Bovine mastitis is a common and costly disease in the dairy industry and is known to negatively affect the amount of epithelium in nonlactating mammary glands. Despite this recognition, an understanding of the mechanisms contributing to reductions in epithelium is lacking. The objective of this study was to evaluate cellular apoptosis and proliferation in uninfected and Staphylococcus aureus-infected mammary glands that were stimulated to rapidly grow and develop. Estradiol and progesterone injections were administered to 18 nonlactating dairy cows to induce mammary growth, and 2 quarters from each animal were infused with saline or Staph. aureus. Mammary tissues were collected at 5 (n = 9) and 10 d (n = 9) postinfusion and examined using quantitative bright field and florescent immunohistochemistry. Staphylococcus aureus mammary glands tended to have a greater number of mammary epithelial cells undergoing apoptosis than saline quarters. In the stromal compartment, challenged quarters contained a lower proportion of cells undergoing apoptosis than saline quarters overall; however, cell types undergoing apoptosis were differentially affected. Staphylococcus aureus quarters contained a lesser percentage of apoptotic fibroblasts while also containing more nonapoptotic immune cells than saline quarters in the intralobular stroma compartment. A similar number of proliferating epithelial cells were present in Staph. aureus and saline mammary tissues, but more proliferating cells were present in the intralobular stroma compartment of Staph. aureus-infused quarters than those infused with saline. When these cellular responses are considered together, it indicates that changes in cellular apoptosis and proliferation contribute to changes in the gland structure by potentiating the expansion of the intralobular stromal compartment, via cellular accumulation, and limiting the amount of epithelium due to increases in cellular apoptosis in affected glands. Reductions in mammary epithelium are expected to reduce future milk yields and productive herd life.

Identification of factors influencing teat dip efficacy trial results by meta-analysis.

Two meta-analyses were conducted using data from peer-reviewed natural exposure (NE) and experimental challenge (EC) teat dip efficacy trials to identify factors influencing the new intramammary infection (IMI) rate. A NE data set containing 16 studies and an EC data set containing 21 studies were created. New IMI rate was calculated based on the percentage of new quarter infections per month (PNQI/mo) for each observation, in both data sets, and used as the dependent variable for model derivation. A linear, mixed-effects model with a random study effect, weighted by number of quarters eligible for infection, was derived for each data set. The final NE model included the effects of experimental design (split herd or split udder), mastitis pathogen group (Staphylococcus aureus, Streptococcus agalactiae, environmental streptococci, gram-negative species, Corynebacterium spp., or coagulase-negative staphylococci), postmilking treatment (iodine, chlorhexidine, linear dodecyl benzene sulfonic acid, chlorine compounds, phenol compounds, or undipped negative controls), and the interaction between mastitis pathogen group and postmilking treatment. Overall, Corynebacterium spp. had the highest new IMI rate (0.0139±0.0018 PNQI/mo), and environmental streptococci and gram-negative species had the lowest (0.0023±0.0022 PNQI/mo). Additionally, trials utilizing a split herd experimental design had a 2-fold higher new IMI rate than trials using a split udder design. The final EC model included the effects of mastitis pathogen (Staph. aureus and Strep. agalactiae), postmilking treatment (iodine, chlorine compounds, "other" active ingredients, or undipped negative controls), geographic region of study (Eastern, Southern, and Pacific Northwest), and the 2-way interactions of region and pathogen group and postmilking treatment and pathogen group. Overall, Staph. aureus and Strep. agalactiae had similar new IMI rates. Quarters dipped postmilking in either iodine (0.0127±0.0099 PNQI/mo), chlorine compounds (0.0258±0.0095 PNQI/mo), or "other" active ingredient teat dips (0.0263±0.0106 PNQI/mo) had lower new IMI rates than undipped quarters (0.0859±0.0087 PNQI/mo). These results indicate that experimental design influences the new IMI rate of teat dip efficacy trials and that using an effective postmilking teat dip has a greater effect on controlling the new Staph. aureus and Strep. agalactiae IMI rate than the teat dip's active ingredient.

Modulation of innate immune function and phenotype in bred dairy heifers during the periparturient period induced by feeding an immunostimulant for 60 days prior to delivery.

The purpose of this study was to evaluate the effect of a feed additive (OmniGen-AF(®), reported to have immune modulating activity) on innate immunity and health events during the periparturient period in dairy heifers when immunity is suppressed. From 60 days prepartum through calving, supplemented heifers (n=20) received OmniGen-AF(®) daily and were compared with unsupplemented controls (n=20). Blood leukocyte innate immune activity (phenotype markers, phagocytic activity, and reactive oxygen species--ROS production) was measured prior to feeding (60 days prepartum), 30 days later, and on days 1, 7, 14, and 30 postpartum. Adverse health events (udder edema, ketosis, displaced abomasum, and death) and milk production were measured at calving and into early lactation. The fraction of leukocytes with measurable CD62L (L-selectin) on their surface from supplemented heifers tended to be greater during the periparturient period in treated heifers than controls (p=0.100). Likewise, leukocyte phagocytosis of Escherichia coli and Staphylococcus aureus during this time period tended to be greater in heifers supplemented with OmniGen-AF(®) (p=0.100). Conversely, ROS production in response to phorbol myristate acetate or when leukocytes were stimulated with killed S. aureus lysate tended to be greater among control heifers compared with supplemented animals (p=0.100). Supplemented heifers exhibited fewer incidents of udder edema than controls (p=0.030) and tended to exhibit a lower rate of new cases of mastitis (p=0.098); however, no differences were observed in milk somatic cell counts or level of milk production. Results demonstrate a positive role of OmniGen-AF(®) in amplifying leukocyte function consistent with antibacterial activity during the periparturient period, and support the continued study of dietary supplementation to enhance mammary gland health in dairy cows.

Use of a staphylococcal vaccine to reduce prevalence of mastitis and lower somatic cell counts in a registered Saanen dairy goat herd.

This investigation evaluated the efficacy of a bacterin in reducing the prevalence of staphylococcal mastitis and somatic cell counts (SCC) in a dairy goat herd. Does were vaccinated or left as controls, and the levels of mastitis and SCC monitored over 18 months. Staphylococcus caprae (42.5%), S. xylosus (15.1%), and S. simulans (10.0%) were the predominant causes of intramammary infections (IMI). The infection rate was 1.64 IMI/doe among vaccinates, which tended to be lower (P < 0.12) than controls (2.67 IMI/doe). The spontaneous cure rate of IMI after immunization was 1.28 cures/doe in vaccinates, which was higher than controls (0.6 cures/doe; P < 0.043). Average SCC of milk samples from vaccinates tended to be lower than that of controls (1274 × 10(3)/ml vs. 1529 × 10(3)/ml, respectively) (P < 0.10). Results support the continued study of mastitis vaccines for use in managing staphylococcal mastitis and SCC in dairy goats.

Effect of dietary supplementation on the antimicrobial activity of blood leukocytes isolated from Holstein heifers.

The purpose of this investigation was to evaluate the effect of an immunostimulating feed supplement (OmniGen-AF®) on the antimicrobial properties of blood leukocytes in dairy heifers in an attempt to prevent mastitis. Blood leukocytes from supplemented and unsupplemented controls were used. Phagocytic activity and reactive oxygen species (ROS) production were studied on d 0 (prior to feed supplementation) and on days 30 and 60 after supplementation. L-selectin and IL-8R mRNA expressions on blood leukocytes were evaluated on d 0 (prior to feed supplementation) and monthly thereafter for 15 mo. On d 30 after supplementation, neutrophils from treated heifers exhibited greater binding and internalization of Escherichia coli and greater ROS production compared with unsupplemented controls. L-selectin mRNA expression was increased in supplemented heifers vs. controls; however, IL-8R mRNA expression was not different. Results support the continued study of dietary supplementation as an additional management tool to enhance udder health in dairy heifers.

Influence of horn flies (Haematobia irritans) on teat skin condition, intramammary infection, and serum anti-S. aureus antibody titres in holstein heifers.

Staphylococcus aureus remains a major mastitis-causing pathogen in growing dairy heifers, resulting in damage to developing milk secretory tissue. The purpose of this study was to evaluate the role of horn flies as vectors in the spread of S. aureus among dairy heifers immunized with a S. aureus bacterin. We analyzed the prevalence of mastitis among quarters, evaluated teat skin condition (as a result of biting flies) prior to and after insecticide administration, and measured serum anti-S. aureus antibody titres monthly after vaccination. Response to S. aureus immunization was poor; however, titres increased 2- to 3-fold during the period when fly populations increased drastically and teat skin condition worsened, especially front quarter teat condition. Presence of flies and the resulting teat lesions were associated with a high level of S. aureus mastitis. Use of an insecticidal pour-on reduced fly populations and healed teat lesions, but existing cases of mastitis required antibiotic therapy.

Technical note: Development of a challenge model for Streptococcus uberis mastitis in dairy heifers.

A challenge model for experimentally inducing Streptococcus uberis mastitis in bred dairy heifers was developed. Qualifying heifers (n=7) exhibited antibody titers of <1:10,000 against Strep. uberis antigens and were free of intramammary infections (IMI). Two contralateral quarters of each heifer were assigned to receive an infusion of Strep. uberis (1,000 to 2,000 cfu); remaining quarters served as unchallenged controls. For a successful challenge and infection, 3 of 4 consecutive mammary secretion samples had to culture positive for Strep. uberis. Six of the 7 heifers were challenged successfully in both infused quarters with a mean dose of 1,080 cfu; once confirmed, infections were treated with a one-time infusion of nonlactating cow therapy. Before challenge, mammary secretion leukocyte counts averaged 8.4×10(6)/mL in all quarters. At 24h after challenge, leukocyte count increased to 18.4×10(6)/mL in challenged quarters, peaking on d 5 at 24.3×10(6)/mL; unchallenged quarters remained at ≤10.4×10(6)/mL, but increased to 15.2×10(6)/mL on d 7 and then decreased. Before challenge, macrophages predominated (81%) in mammary secretions followed by lymphocytes (15.3%) and neutrophils (3.7%). By 24h after challenge, neutrophils increased in challenged quarters and predominated for the duration of the trial (65.3 to 70%), whereas macrophages predominated in unchallenged control quarters (65.2 to 75.2%). The challenge model was successful in establishing Strep. uberis IMI in 85.7% of animals, and IMI were controlled (100% cure) by administering nonlactating cow therapy. All heifers calved free of IMI and antimicrobial residues, with milk production similar to that of herd mates and with somatic cell counts (SCC) <200,000 cells/mL.

Curli production and genetic relationships among Escherichia coli from cases of bovine mastitis.

Curli are adhesive surface structures produced by some Escherichia coli and Salmonella strains that bind host proteins and activate inflammatory mediators. In this study, 61 E. coli isolates from 36 clinical cases of bovine mastitis were characterized using enterobacterial repetitive intergenic consensus-PCR and screened for their ability to produce curli. Effect of curli production on case recovery, based on a return to precase milk yield, was investigated for a subset of 43 isolates from 20 quarters of 19 cows. Thirty-five (57%) of 61 isolates were curli positive. Fifty-eight of the 61 isolates clustered into 2 clonal groups at 52% genetic similarity. Genetically diverse E. coli isolates were simultaneously cultured from individual cases. Twenty-three isolates from 13 cows were clustered in clonal group I, of which 5 cases (38%) were curli positive; 35 isolates from 22 cows were clustered in clonal group II, of which 15 cases (68%) were curli positive. No association was found between genetic similarity and phenotypic curli expression of isolates from cows with clinical E. coli mastitis cases. Phenotypic curli expression in isolates did not affect recovery of cows' milk yield to premastitis production levels.

Germicidal activities of representatives of five different teat dip classes against three bovine mycoplasma species using a modified excised teat model.

Six representative teat dips from five different teat dip classes were tested for germicidal activity against challenge exposure to Mycoplasma bovis, Mycoplasma californicum, and Mycoplasma bovigenitalium using a modified excised teat model. All teat dip formulations tested were efficacious against all of the Mycoplasma species, providing bacterial logarithmic reductions above 4. The germicides performed best against M. bovigenitalium with an average log reduction (LR) of 6.29. Average LR were 5.41 and 5.70 against M. bovis and M. californicum, respectively. The iodine and chlorhexidine products performed best against M. bovis and M. californicum, respectively, with complete kill of all organisms. The chlorhexidine and the barrier chlorine product also had complete kill of M. bovigenitalium organisms.

Reduction of mastitis caused by experimental challenge with Staphylococcus aureus and Streptococcus agalactiae by use of a quaternary ammonium and halogen-mixture teat dip.

A teat-dip formulation containing sodium dichloro isocyanuric acid, bronopol, and quaternary ammonium was tested for efficacy against Staphylococcus aureus and Streptococcus agalactiae intramammary infections (IMI) using an experimental challenge model. Sixty-two Jersey cows from the Hill Farm Research Station (Homer, LA) were used in an 8-wk controlled infection trial to evaluate the teat dip. During the afternoon milking, Monday through Friday for 8 wk, all teats of each cow were immersed to a depth of approximately 25 mm in a challenge suspension containing approximately 5 x 10(7) cfu of Staphylococcus aureus and approximately 5 x 10(7) cfu of Streptococcus agalactiae immediately after milking machines were removed. Immediately after challenge, the distal 25 mm of two contralateral teats were dipped with the experimental teat dip; the remaining two teats served as undipped controls. The experimental teat dip reduced the number of new Staph. aureus IMI by 70.9% and reduced the number of new Strep. agalactiae IMI by 60.0%. Teat end and teat skin condition were characterized as normal and without irritation at the completion of the study. The combination of the three germicides in this experimental teat dip is unique and an effective formulation without adverse effects on condition of teat ends or teat skin.

Prevalence of mastitis in dairy heifers and effectiveness of antibiotic therapy.

Dairy heifers were treated 0 to 90 d, 90 to 180 d, or 180 to 270 d prepartum with one of five different antibiotic products to determine the best time and with which product they should be treated prior to calving. Two hundred thirty-three heifers were included in the study. At the initial sampling, 56.5% of quarters were infected with some type of organism and 15.4% of quarters were infected with Staphylococcus aureus. Treatments included a cephapirin dry cow product, a penicillin-novobiocin dry cow product, a penicillin-streptomycin dry cow product, an experimental dry cow product containing tilmicosin, and a cephalonium dry cow product not available in the United States. Cure rates for the five antibiotic products indicated that all were equally effective against Staph. aureus and all were significantly more effective than the spontaneous cure rate observed in untreated control quarters. No differences in efficacy were observed due to the different treatment times prepartum. However, fewer new Staph. aureus infections occurred after treatment in the group treated at 180 to 270 d prepartum, indicating that treatment in the third trimester will reduce the chances of new intramammary infections occurring after treatment and persisting to calving.

A comparison of two commercially available Escherichia coli J5 vaccines against E. coli intramammary challenge.

The efficacy of two commercially available Escherichia coli J5 bacterins was investigated. Jersey cows were randomly assigned to one of three treatment groups: 1) unvaccinated controls, 2) vaccinated with J.VAC (Merial Limited, Athens, GA), and 3) vaccinated with J5 bacterin. All cows were vaccinated at drying off and at 2 wk before anticipated calving. Cows that were vaccinated with the J5 bacterin also received a third immunization at calving. One quarter of each cow was challenged with approximately 64 cfu of E. coli at 14 to 30 d postcalving. Immunization by either vaccine did not influence the severity of coliform mastitis; however, the mean number of colony-forming units of E. coli recovered from challenged quarters was significantly lower for immunized cows than for control cows at 144 h postchallenge. Serum and mammary secretion immunoglobulin (Ig)G, IgG1, and IgG2 titers against E. coli J5 whole-cell antigens were enhanced in vaccinated cows. Serum and mammary secretion IgM were not different among treatment groups. Somatic cell counts in milk from challenged quarters, rectal temperatures, and the clinical status of cows following intramammary challenge were not different among treatment groups.

Efficacies of chlorine dioxide and lodophor teat dips during experimental challenge with Staphylococcus aureus and Streptococcus agalactiae.

We tested two postmilking teat dips for efficacy against Staphylococcus aureus and Streptococcus agalactiae using experimental challenge procedures recommended by the National Mastitis Council. The chlorine dioxide teat dip that contained 0.7% sodium chlorite reduced the number of new intramammary infections (IMI) caused by Staph. aureus by 86.6% and reduced new IMI caused by Strep. agalactiae by 88.4%. The 0.5% iodophor teat dip reduced the number of new IMI caused by Staph. aureus by 92.9% and reduced the number of new IMI caused by Strep. agalactiae by 43.4%. Teat skin and teat end conditions were evaluated before and after the study, and no deleterious effects were noted among dipped quarters compared with undipped control quarters for either teat dip.

Deoxyribonucleic acid fingerprinting of Staphylococcus aureus from heifer mammary secretions and from horn flies.

Staphylococcus aureus isolated from heifer mammary secretions, streak canals, and horn flies (Haematobia irritans) were evaluated by randomly amplified polymorphic DNA fingerprinting. The relationship between DNA fingerprint patterns of S. aureus isolated from horn flies and S. aureus isolated from heifer mammary glands was examined. Amplified DNA fragments were visualized by agarose gel electrophoresis and were analyzed by densitometry. Analysis of DNA fingerprint patterns of 56 S. aureus isolates that were obtained from heifer mammary secretions or streak canals resulted in three distinct subtypes of S. aureus. Of these, 31 isolates (55%) belonged to subtype 1, 22 isolates (39%) belonged to subtype 2, and 3 (5%) belonged to subtype 3. Eight of 10 S. aureus isolates from horn flies belonged to subtype 1, and 2 isolates belonged to subtype 2. Thus, all of the S. aureus isolates from horn flies had DNA fingerprint patterns identical to the majority (95%) of S. aureus isolates from heifer mammary secretions or streak canals. In addition, 10 S. aureus isolates from multiparous cows from the same herd were examined by randomly amplified polymorphic DNA. All S. aureus isolates from multiparous cows belonged to subtype 3. Results of this study suggest that horn flies may play an important role in the transmission of S. aureus to nulligravid and primigravid heifers. Furthermore, this study demonstrates the usefulness of randomly amplified polymorphic DNA fingerprinting to distinguish between different subtypes of S. aureus and to draw epidemiological inferences from the information it provides.

Efficacy of parenterally or intramammarily administered tilmicosin or ceftiofur against Staphylococcus aureus mastitis during lactation.

Two antibiotic preparations, tilmicosin and ceftiofur, were tested intramammarily and parenterally against Staphylococcus aureus mastitis in lactating cows. Neither product was effective as a lactating cow treatment at the doses and durations of treatment tested. Injection or infusion of tilmicosin and infusion of ceftiofur resulted in reductions of bacteria present in milk; however, only one quarter treated with infusion of tilmicosin was cured, and no cures were observed for the other treatments. Somatic cell counts were transiently reduced by infusion of ceftiofur and by infusion and injection of tilmicosin; however, they returned to pretreatment values by 28 d posttreatment.

Comparison of tilmicosin and cephapirin as therapeutics for Staphylococcus aureus mastitis at dry-off.

Forty-four cows (26 Jerseys and 18 Holsteins) that had at least 1 mammary quarter that was naturally (n = 12) or experimentally (n = 84) infected with Staphylococcus aureus were allotted to three treatment groups of approximately equal number at the end of lactation. Cows were dried off by abrupt cessation of milking, and dry cow therapy was administered as an intramammary infusion of cephapirin benzathine at 10 ml per quarter, an intramammary infusion of tilmicosin (solution containing 300 mg/ml) at 5 ml per quarter, or a subcutaneous injection of tilmicosin at 5 mg/kg of body weight on the day of drying off and another injection 4 d later. Mammary secretions were monitored during the dry period and postpartum for antimicrobial residues, intramammary infection (IMI) status, and somatic cell counts. Results demonstrated the following percentage cures for IMI caused by Staph. aureus at 28 d postcalving based on individual mammary quarters: cephapirin benzathine, 78.1%; tilmicosin infused, 74.2%; and tilmicosin injected, 9.1%. During the first 4 wk after drying off, the mean concentration of tilmicosin in mammary secretions from cows infused with the antibiotic remained approximately 10-fold higher than that in secretions from cows injected with the antibiotic (3.43 vs. 0.32 ppm), and, by the time of calving, concentrations for cows treated with both methods were below the dilution limit of the assay (< 0.1 ppm). Results demonstrated that intramammary infusion of tilmicosin was equally as effective as cephapirin benzathine in curing IMI caused by Staph. aureus at drying off; however, the subcutaneous injection of tilmicosin at the dose used was not effective as a dry cow therapeutic against Staph. aureus.